Structural dynamics in the C-terminal domain of calmodulin at low calcium levels

A. Malmendal, J. Evenäs, S. Forsén, M. Akke

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

Resumé

Calmodulin undergoes Ca2+-induced structural rearrangements that are intimately coupled to the regulation of numerous cellular processes. The C-terminal domain of calmodulin has previously been observed to exhibit conformational exchange in the absence of Ca2+. Here, we characterize further the conformational dynamics in the presence of low concentrations of Ca2+using15N spin relaxation experiments. The analysis included1H-15N dipolar/15N chemical shift anisotropy interference cross-correlation relaxation rates to improve the description of the exchange processes, as well as the picosecond to nanosecond dynamics. Conformational transitions on microsecond to millisecond time scales were revealed by exchange contributions to the transverse auto-relaxation rates. In order to separate the effects of Ca2+exchange from intramolecular conformational exchange processes in the apo state, transverse autorelaxation rates were measured at different concentrations of free Ca2+. The results reveal a Ca2+-dependent contribution due mainly to exchange between the apo and (Ca2+1, states with an apparent Ca2+off-rate of 5115 s-1, as well as Ca2+-independent contributions due to conformational exchange within the apo state.15N chemical shift differences estimated from the exchange data suggest that the first Ca2+binds preferentially to loop IV. Thus, characterization of chemical exchange as a function of Ca2+concentration has enabled the extraction of unique information on the rapidly exchanging and weakly populated (textless 10%) (Ca2+)1state that is otherwise inaccessible to direct study due to strongly cooperative Ca2+binding. The conformational exchange within the apo state appears to involve transitions between a predominantly populated closed conformation and a smaller population of more open conformations. The picosecond to nanosecond dynamics of the apo state are typical of a well-folded protein, with reduced amplitudes of motions in the helical segments, but with significant flexibility in the Ca2+-binding loops. Comparisons with older parameters for skeletal troponin C and calbindin D(9k) reveal key structural and dynamical differences that correlate with the different Ca2+-binding properties of these proteins.
OriginalsprogEngelsk
TidsskriftJournal of Molecular Biology
Vol/bind293
Udgave nummer4
ISSN0022-2836
DOI
StatusUdgivet - 1999

Emneord

  • Calcium binding
  • Calmodulin
  • Conformational exchange
  • NMR
  • Protein dynamics

Citer dette

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title = "Structural dynamics in the C-terminal domain of calmodulin at low calcium levels",
abstract = "Calmodulin undergoes Ca2+-induced structural rearrangements that are intimately coupled to the regulation of numerous cellular processes. The C-terminal domain of calmodulin has previously been observed to exhibit conformational exchange in the absence of Ca2+. Here, we characterize further the conformational dynamics in the presence of low concentrations of Ca2+using15N spin relaxation experiments. The analysis included1H-15N dipolar/15N chemical shift anisotropy interference cross-correlation relaxation rates to improve the description of the exchange processes, as well as the picosecond to nanosecond dynamics. Conformational transitions on microsecond to millisecond time scales were revealed by exchange contributions to the transverse auto-relaxation rates. In order to separate the effects of Ca2+exchange from intramolecular conformational exchange processes in the apo state, transverse autorelaxation rates were measured at different concentrations of free Ca2+. The results reveal a Ca2+-dependent contribution due mainly to exchange between the apo and (Ca2+1, states with an apparent Ca2+off-rate of 5115 s-1, as well as Ca2+-independent contributions due to conformational exchange within the apo state.15N chemical shift differences estimated from the exchange data suggest that the first Ca2+binds preferentially to loop IV. Thus, characterization of chemical exchange as a function of Ca2+concentration has enabled the extraction of unique information on the rapidly exchanging and weakly populated (textless 10{\%}) (Ca2+)1state that is otherwise inaccessible to direct study due to strongly cooperative Ca2+binding. The conformational exchange within the apo state appears to involve transitions between a predominantly populated closed conformation and a smaller population of more open conformations. The picosecond to nanosecond dynamics of the apo state are typical of a well-folded protein, with reduced amplitudes of motions in the helical segments, but with significant flexibility in the Ca2+-binding loops. Comparisons with older parameters for skeletal troponin C and calbindin D(9k) reveal key structural and dynamical differences that correlate with the different Ca2+-binding properties of these proteins.",
keywords = "Calcium binding, Calmodulin, Conformational exchange, NMR, Protein dynamics",
author = "A. Malmendal and J. Even{\"a}s and S. Fors{\'e}n and M. Akke",
year = "1999",
doi = "10.1006/jmbi.1999.3188",
language = "English",
volume = "293",
journal = "Journal of Molecular Biology",
issn = "0022-2836",
publisher = "Academic Press",
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Structural dynamics in the C-terminal domain of calmodulin at low calcium levels. / Malmendal, A.; Evenäs, J.; Forsén, S.; Akke, M.

I: Journal of Molecular Biology, Bind 293, Nr. 4, 1999.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

TY - JOUR

T1 - Structural dynamics in the C-terminal domain of calmodulin at low calcium levels

AU - Malmendal, A.

AU - Evenäs, J.

AU - Forsén, S.

AU - Akke, M.

PY - 1999

Y1 - 1999

N2 - Calmodulin undergoes Ca2+-induced structural rearrangements that are intimately coupled to the regulation of numerous cellular processes. The C-terminal domain of calmodulin has previously been observed to exhibit conformational exchange in the absence of Ca2+. Here, we characterize further the conformational dynamics in the presence of low concentrations of Ca2+using15N spin relaxation experiments. The analysis included1H-15N dipolar/15N chemical shift anisotropy interference cross-correlation relaxation rates to improve the description of the exchange processes, as well as the picosecond to nanosecond dynamics. Conformational transitions on microsecond to millisecond time scales were revealed by exchange contributions to the transverse auto-relaxation rates. In order to separate the effects of Ca2+exchange from intramolecular conformational exchange processes in the apo state, transverse autorelaxation rates were measured at different concentrations of free Ca2+. The results reveal a Ca2+-dependent contribution due mainly to exchange between the apo and (Ca2+1, states with an apparent Ca2+off-rate of 5115 s-1, as well as Ca2+-independent contributions due to conformational exchange within the apo state.15N chemical shift differences estimated from the exchange data suggest that the first Ca2+binds preferentially to loop IV. Thus, characterization of chemical exchange as a function of Ca2+concentration has enabled the extraction of unique information on the rapidly exchanging and weakly populated (textless 10%) (Ca2+)1state that is otherwise inaccessible to direct study due to strongly cooperative Ca2+binding. The conformational exchange within the apo state appears to involve transitions between a predominantly populated closed conformation and a smaller population of more open conformations. The picosecond to nanosecond dynamics of the apo state are typical of a well-folded protein, with reduced amplitudes of motions in the helical segments, but with significant flexibility in the Ca2+-binding loops. Comparisons with older parameters for skeletal troponin C and calbindin D(9k) reveal key structural and dynamical differences that correlate with the different Ca2+-binding properties of these proteins.

AB - Calmodulin undergoes Ca2+-induced structural rearrangements that are intimately coupled to the regulation of numerous cellular processes. The C-terminal domain of calmodulin has previously been observed to exhibit conformational exchange in the absence of Ca2+. Here, we characterize further the conformational dynamics in the presence of low concentrations of Ca2+using15N spin relaxation experiments. The analysis included1H-15N dipolar/15N chemical shift anisotropy interference cross-correlation relaxation rates to improve the description of the exchange processes, as well as the picosecond to nanosecond dynamics. Conformational transitions on microsecond to millisecond time scales were revealed by exchange contributions to the transverse auto-relaxation rates. In order to separate the effects of Ca2+exchange from intramolecular conformational exchange processes in the apo state, transverse autorelaxation rates were measured at different concentrations of free Ca2+. The results reveal a Ca2+-dependent contribution due mainly to exchange between the apo and (Ca2+1, states with an apparent Ca2+off-rate of 5115 s-1, as well as Ca2+-independent contributions due to conformational exchange within the apo state.15N chemical shift differences estimated from the exchange data suggest that the first Ca2+binds preferentially to loop IV. Thus, characterization of chemical exchange as a function of Ca2+concentration has enabled the extraction of unique information on the rapidly exchanging and weakly populated (textless 10%) (Ca2+)1state that is otherwise inaccessible to direct study due to strongly cooperative Ca2+binding. The conformational exchange within the apo state appears to involve transitions between a predominantly populated closed conformation and a smaller population of more open conformations. The picosecond to nanosecond dynamics of the apo state are typical of a well-folded protein, with reduced amplitudes of motions in the helical segments, but with significant flexibility in the Ca2+-binding loops. Comparisons with older parameters for skeletal troponin C and calbindin D(9k) reveal key structural and dynamical differences that correlate with the different Ca2+-binding properties of these proteins.

KW - Calcium binding

KW - Calmodulin

KW - Conformational exchange

KW - NMR

KW - Protein dynamics

U2 - 10.1006/jmbi.1999.3188

DO - 10.1006/jmbi.1999.3188

M3 - Journal article

VL - 293

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 4

ER -